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Pretargeted nuclear imaging based on the ligation between tetrazines and nano-sized targeting agents functionalized with trans-cyclooctene (TCO) has recently been shown to improve both imaging contrast and dosimetry in nuclear imaging of nanomedicines. Herein, we describe the improved radiosynthesis of a 11 C-labeled tetrazine ([ 11 C]AE-1) and its preliminary evaluation in both mice and pigs. Pretargeted imaging in mice was carried out using both a new TCO-functionalized polyglutamic acid and a previously reported TCO-functionalized bisphosphonate system as targeting agents. Unfortunately, pretargeted imaging was not successful using these targeting agents in pair with [ 11 C]AE-1. However, brain imaging in pig indicated that the tracer crossed the blood-brain-barrier. Hence, we suggest that this tetrazine scaffold could be used as a starting point for the development of pretargeted brain imaging, which has so far been a challenging task.

Pretargeted nuclear imaging and radiotherapy have recently attracted increasing attention for diagnosis and treatment of cancer with nanomedicines. This is because it conceptually offers better imaging contrast and therapeutic efficiency while reducing the dose to radiosensitive tissues compared to conventional strategies. In conventional imaging and radiotherapy, a directly radiolabeled nano-sized vector is administered and allowed to accumulate in the tumor, typically on a timescale of several days. In contrast, pretargeting is based on a two-step approach. First, a tumor-accumulating vector carrying a tag is administered followed by injection of a fast clearing radiolabeled agent that rapidly recognizes the tag of the tumor-bound vector in vivo. Therefore, pretargeting circumvents the use of long-lived radionuclides that is a necessity for sufficient tumor accumulation and target-to-background ratios using conventional approaches. In this review, we give an overview of recent advances in pretargeted imaging strategies. We will critically reflect on the advantages and disadvantages of current state-of-the-art conventional imaging approaches and compare them to pretargeted strategies. We will discuss the pretargeted imaging concept and the involved chemistry. Finally, we will discuss the steps forward in respect to clinical translation, and how pretargeted strategies could be applied to improve state-of-the-art radiotherapeutic approaches.

Plasmonic nanoparticle-based photothermal cancer therapy is a promising new tool to inflict localized and irreversible damage to tumor tissue by hyperthermia, without harming surrounding healthy tissue. We developed a single particle and positron emission tomography (PET)-based platform to quantitatively correlate the heat generation of plasmonic nanoparticles with their potential as cancer killing agents. In vitro, the heat generation and absorption cross-section of single irradiated nanoparticles were quantified using a temperature sensitive lipid-based assay and compared to their theoretically predicted photo-absorption. In vivo, the heat generation of irradiated nanoparticles was evaluated in human tumor xenografts in mice using 2-deoxy-2-[F-18]fluoro-D-glucose ((18)F-FDG) PET imaging. To validate the use of this platform, we quantified the photothermal efficiency of near infrared resonant silica-gold nanoshells (AuNSs) and benchmarked this against the heating of colloidal spherical, solid gold nanoparticles (AuNPs). As expected, both in vitro and in vivo the heat generation of the resonant AuNSs performed superior compared to the non-resonant AuNPs. Furthermore, the results showed that PET imaging could be reliably used to monitor early treatment response of photothermal treatment. This multidisciplinary approach provides a much needed platform to benchmark the emerging plethora of novel plasmonic nanoparticles for their potential for photothermal cancer therapy.

Upon infection by bacteriophage λ the λ repressor protein, CI, interacts with the λ operator DNA, thereby regulating protein expression and deciding between the lysogenic and lytic state. This λ switch is a model on the basis of which epigenetic switch regulation is understood. In order to study the interaction between naturally supercoiled DNA and the DNA associating protein CI, we invented a novel assay where supercoiled circular DNA plasmids were individually tethered by peptide nucleic acid (PNA) handles [1]. We used this tethered plasmid assay for a single molecule investigation of the dynamics of supercoiled DNA and studied both the dynamics of the molecule itself and its interactions with the regulatory CI protein. The dynamics of the construct was analyzed by tracking the tethered bead. This revealed that compared with relaxed DNA, the presence of supercoils greatly enhances juxtaposition probability [2]. When CI was added to the supercoiled assay, the protein would attach to the operator sites thereby looping DNA. Our studies reveal that the efficiency and cooperativity of the epigenetic λ switch are significantly increased in the supercoiled system compared with a linear assay, thus increasing the Hill coefficient [2,3]. In contrast to other single molecule assays, the current methodology allows for studying DNA dynamics and DNA-protein interactions of DNA in its naturally supercoiled state.

Hyperthermia has great potential as a cancer therapy as it weakens or causes irreversible damage to cancer cells. However, available heat sources are poor in discriminating between healthy and cancerous tissue. In this thesis work, the application of plasmonic nanoparticles as photo-induced strong, localized thermal transducers was investigated for cancer therapy. Gold nanoparticles exhibit surface plasmon resonance that greatly enhances their photoabsorption properties. When irradiated with resonant light, they eciently absorb the light and convert it into extremely local and well-controlled heating with temperature increases that easily exceed 100. Due to these unique optical properties and their biocompatibility, gold nanoparticles are promising candidates for selective photothermal cancer therapy. Light with wavelengths in the near-infrared (NIR) region has low absorption and high penetration through biological material. Hence, the combination of these two non-destructive moieties can inict irreversible damage to the tumor tissue by strong localized hyperthermia, without harming the surrounding healthy tissue. However in the literature, the optimal choice of plasmonic nanoparticles for this therapy remains an open question. Using positron emission tomography/computed tomography (PET/CT) imaging as a treatment evaluation tool it was found that NIR irradiated resonant silica-gold nanoshells had a higher therapeutic ecacy than non-resonant colloidal gold nanoparticles, when delivered directly into subcutaneous tumor xenografts in mice. To better understand the photo-physical properties, the plasmonic heating of the resonant and non-resonant nanoparticles was also compared using an in vitro temperature sensitive assay. This assay enabled measurements of the heat generation of single NIR irradiated nanoparticles and con rmed that the resonant silica-gold nanoshells were superior to the non-resonant nanoparticles. These ndings were in agreement with